Although the glomerulus has long since been known to be compromised of four resident cell types, the contributions of the endothelial cell, mesangial cell, and podocyte to glomerular architecture and function have garnered virtually all of the research attention to date. In contrast, very little is known about the normal biological function of parietal epithelial cells (PECs), and how these cells respond to injury in disease states. Despite originating from the same mesenchymal cell as podocytes, mature PECs represent a distinctly unique cell type as they constitutively express different genes, and readily proliferate when injured, contrasting from the typically quiescent podocyte. The overall goals of this grant proposal are to provide novel insights into the biology and function of PECs in health and disease. In the first aim, we will test the hypothesis that PECs have a critical role in the handling of albumin in the glomerular ultrafiltrate. We propose that PECs lining Bowman's basement membrane form a second glomerular permeability barrier, distinct from the conventional glomerular filtration barrier (which comprises the endothelium, GBM, podocytes). We will test the hypothesis that the well-defined tight junctions and junctional proteins between neighboring cells are abnormal in experimental disease causing increased albumin permeability into the peri-glomerular space. Studies will be conducted in cultured PECs and in experimental models of podocyte and PEC disease in Ksp-EYFP mice (PECs are green) injected with different types and sizes of fluorescent molecular probes as tracers. We will also test the hypothesis that PECs normally take up filtered albumin into vacuoles for lysozymal degradation. However, when proteinuria is marked in primary podocyte disease, we propose that increased uptake leading to excess intra-PEC albumin content is then injurious. The second aim tests the hypothesis that a critical function of PECs is to serve as local precursor cells that transform/differentiate into podocytes when podocyte number decreases. We have crossed Ksp-EYFP mice (green PECs) with nephrin CFP mice (blue podocytes). Podocyte apoptosis and thus loss will be induced in two mouse models (diphtheria-toxin receptor specifically in podocytes;anti-podocyte antibody). The migration of fluorescent cells (PECs) will be carefully monitored in a serial fashion and trans-differentiation into podocytes established based on dual expression of green fluorescent tag concurrently with podocyte-specific proteins such as WT-1 and synaptopodin. Concurrent PEC cell culture studies will augment the in vivo experiments. In summary, these studies are aimed at providing important and novel information into the normal function of PECs, and how PECs respond to direct injury and to injury of their neighboring podocytes. PUBLIC HEALTH RELEVANCE: Kidney disease is increasing at epidemic rates in the U.S. One out of ten adults has some form of kidney damage. The overall goal of this grant is to delineate new paradigms in the biology of parietal epithelial cells, so that ultimately new strategies can be developed to reduce the burden of kidney disease